Abstract Systemic lupus erythematosus (SLE) is a prototypic autoimmune disease characterized by the production of antinuclear antibodies (ANA) in association with multisystem inflammatory manifestations. These antibodies target a wide array of nuclear macromolecules and mediate disease in part by the formation of immune complexes that promote inflammation and tissue damage. While antibodies to nucleosomes and its DNA and histone components are the most characteristic of SLE, antibodies to RNA and RNA binding proteins may display similar pathogenetic properties. As shown in in vivo and in vitro experiments, nuclear antigens, including DNA, RNA and the high mobility group protein 1 (HMGB1), have activity as alarmins. Alarmins represent endogenous molecules that can be released from activated as well as dying cells and can stimulate innate immunity. Once they translocate into the extracellular milieu, nuclear antigens can stimulate toll-like receptors (TLRs), non-TLR nucleic acid sensors as well as receptors binding other components of the immune complexes. Because of their alarmin activity, nuclear molecules can play at least 3 roles in disease: 1) drive the production of antinuclear antibodies by autoantigen-specific B and T cells; 2) function as autoadjuvants to stimulate immune cell activation; and 3) form immune complexes that deposit in the tissue and incite inflammation. In view of these roles, the release or exposure of nuclear antigens in an immunologically relevant form is a key step in pathogenesis. To elucidate the release of nuclear antigens from cells, their physical-chemical properties and their signaling activity, we propose investigating these issues in in vivo and in vitro models. Specifically, we will explore the role of nuclear molecules as autoadjuvants and autoantigens in the context of microparticles. Microparticles are small membrane-bound vesicles which, like alarmins, are released from activated or dying cells and display immunostimulatory activity. Three specific aims are proposed: 1) To elucidate the in vivo and in vitro release of DNA, RNA and HMGB1 during activation and cell death, determining the relative distribution in soluble and particulate form; 2) To assess the antigenic properties of microparticles and their ability to form immune complexes in vitro and in vivo in autoimmune mice. These studies will determine whether complexes containing nucleic acids are comprised of microparticles; and 3) To define the immunological activity of microparticles, determining the contribution of DNA and RNA to their ability to stimulate macrophages and dendritic cells. These experiments will also explore novel cationic polymers to block these responses. Successful completion of these experiments will provide new insights in the pathogenic role in SLE of nuclear molecules in the form of microparticles as well as provide new biomarkers and potential targets of therapy. PUBLIC HEALTH RELEVANCE: Project Narrative These studies are relevant to the health of veterans by addressing key issues in the pathogenesis of autoimmune disease. Like other autoimmune diseases, SLE is growing in frequency as the number of women in the military increases. By elucidating the role of extracellular nuclear molecules in forming pathogenic immune complexes, these studies will provide new insights into the mechanisms of inflammation and autoreactivity in SLE. These studies could suggest new biomarkers for autoimmune diseases based on assessing microparticle number and phenotype. Finally, our studies on the immune properties of microparticles could identify new signaling pathways operating in autoimmunity, allowing the development of more effective immunosuppressive agents to improve outcomes and reduce the burden of health care in the veteran population.